Expression of the S1 and S2 subunits of murine coronavirus JHMV spike protein by a vaccinia virus transient expression system.
Identifieur interne : 003535 ( PubMed/Corpus ); précédent : 003534; suivant : 003536Expression of the S1 and S2 subunits of murine coronavirus JHMV spike protein by a vaccinia virus transient expression system.
Auteurs : H. Kubo ; F. TaguchiSource :
- The Journal of general virology [ 0022-1317 ] ; 1993.
English descriptors
- KwdEn :
- Amino Acid Sequence, Animals, Antigens, Viral (analysis), Antigens, Viral (physiology), Base Sequence, Blotting, Western, Cell Line, Genetic Vectors, Membrane Glycoproteins, Mice, Molecular Sequence Data, Murine hepatitis virus (chemistry), Murine hepatitis virus (genetics), Murine hepatitis virus (physiology), Recombinant Proteins (biosynthesis), Spike Glycoprotein, Coronavirus, Vaccinia virus, Viral Envelope Proteins (biosynthesis), Viral Envelope Proteins (genetics), Viral Envelope Proteins (physiology).
- MESH :
- chemical , analysis : Antigens, Viral.
- chemical , biosynthesis : Recombinant Proteins, Viral Envelope Proteins.
- chemical , genetics : Viral Envelope Proteins.
- chemical , physiology : Antigens, Viral, Viral Envelope Proteins.
- chemistry : Murine hepatitis virus.
- genetics : Murine hepatitis virus.
- physiology : Murine hepatitis virus.
- Amino Acid Sequence, Animals, Base Sequence, Blotting, Western, Cell Line, Genetic Vectors, Membrane Glycoproteins, Mice, Molecular Sequence Data, Spike Glycoprotein, Coronavirus, Vaccinia virus.
Abstract
The spike (S) protein of murine coronavirus JHMV, variant cl-2, comprises two polypeptides, N-terminal S1 (with an N-terminal signal peptide) and C-terminal S2 (with a C-terminal transmembrane domain). In order to express these subunits, we constructed three different vaccinia virus transfer vectors (VV-TVs) containing cDNAs encoding the S1 protein without a transmembrane domain (pSFS1utt), the S1 protein with a C-terminal transmembrane domain derived from S2 (pSFS1tmd) or the S2 protein with an N-terminal signal peptide derived from S1 (pSFssS2). The S1 and S2 proteins were expressed in DBT cells by infection with vaccinia virus and transfection of these VV-TVs. In cells transfected with the pSFS1utt and pSFS1tmd, 96K and 106K proteins, respectively, were detected by Western blotting. The ssS2 protein expressed by pSFssS2 was 96K, which was slightly larger than the authentic S2 protein. The S1utt and S1tmd proteins were shown by binding studies using a panel of monoclonal antibodies to be antigenically indistinguishable from the authentic S1 protein. The S1tmd and ssS2 proteins were detected on the cell surface by immunofluorescence, whereas the S1utt protein was not. However, when the S1utt protein was expressed together with the ssS2 protein, the S1utt was detected on the cell membrane. This suggested that the S1utt was associated with ssS2 on the cell membrane. These observations indicate that the expressed S1 and S2 proteins associated in a similar manner to the authentic S1 and S2 proteins produced in DBT cells infected with cl-2. However, cell fusion was not observed in cells expressing either S1 or S2 nor in cells coexpressing both S1 and S2, although the whole S protein expressed by VV-TV did induce fusion.
DOI: 10.1099/0022-1317-74-11-2373
PubMed: 8245853
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pubmed:8245853Le document en format XML
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Taguchi</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="1993">1993</date><idno type="RBID">pubmed:8245853</idno><idno type="pmid">8245853</idno><idno type="doi">10.1099/0022-1317-74-11-2373</idno><idno type="wicri:Area/PubMed/Corpus">003535</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">003535</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Expression of the S1 and S2 subunits of murine coronavirus JHMV spike protein by a vaccinia virus transient expression system.</title><author><name sortKey="Kubo, H" sort="Kubo, H" uniqKey="Kubo H" first="H" last="Kubo">H. Kubo</name><affiliation><nlm:affiliation>National Institute of Neuroscience, NCNP, Tokyo, Japan.</nlm:affiliation></affiliation></author><author><name sortKey="Taguchi, F" sort="Taguchi, F" uniqKey="Taguchi F" first="F" last="Taguchi">F. Taguchi</name></author></analytic><series><title level="j">The Journal of general virology</title><idno type="ISSN">0022-1317</idno><imprint><date when="1993" type="published">1993</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Sequence</term><term>Animals</term><term>Antigens, Viral (analysis)</term><term>Antigens, Viral (physiology)</term><term>Base Sequence</term><term>Blotting, Western</term><term>Cell Line</term><term>Genetic Vectors</term><term>Membrane Glycoproteins</term><term>Mice</term><term>Molecular Sequence Data</term><term>Murine hepatitis virus (chemistry)</term><term>Murine hepatitis virus (genetics)</term><term>Murine hepatitis virus (physiology)</term><term>Recombinant Proteins (biosynthesis)</term><term>Spike Glycoprotein, Coronavirus</term><term>Vaccinia virus</term><term>Viral Envelope Proteins (biosynthesis)</term><term>Viral Envelope Proteins (genetics)</term><term>Viral Envelope Proteins (physiology)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="analysis" xml:lang="en"><term>Antigens, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Recombinant Proteins</term><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="physiology" xml:lang="en"><term>Antigens, Viral</term><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" qualifier="chemistry" xml:lang="en"><term>Murine hepatitis virus</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Murine hepatitis virus</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Murine hepatitis virus</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Animals</term><term>Base Sequence</term><term>Blotting, Western</term><term>Cell Line</term><term>Genetic Vectors</term><term>Membrane Glycoproteins</term><term>Mice</term><term>Molecular Sequence Data</term><term>Spike Glycoprotein, Coronavirus</term><term>Vaccinia virus</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The spike (S) protein of murine coronavirus JHMV, variant cl-2, comprises two polypeptides, N-terminal S1 (with an N-terminal signal peptide) and C-terminal S2 (with a C-terminal transmembrane domain). In order to express these subunits, we constructed three different vaccinia virus transfer vectors (VV-TVs) containing cDNAs encoding the S1 protein without a transmembrane domain (pSFS1utt), the S1 protein with a C-terminal transmembrane domain derived from S2 (pSFS1tmd) or the S2 protein with an N-terminal signal peptide derived from S1 (pSFssS2). The S1 and S2 proteins were expressed in DBT cells by infection with vaccinia virus and transfection of these VV-TVs. In cells transfected with the pSFS1utt and pSFS1tmd, 96K and 106K proteins, respectively, were detected by Western blotting. The ssS2 protein expressed by pSFssS2 was 96K, which was slightly larger than the authentic S2 protein. The S1utt and S1tmd proteins were shown by binding studies using a panel of monoclonal antibodies to be antigenically indistinguishable from the authentic S1 protein. The S1tmd and ssS2 proteins were detected on the cell surface by immunofluorescence, whereas the S1utt protein was not. However, when the S1utt protein was expressed together with the ssS2 protein, the S1utt was detected on the cell membrane. This suggested that the S1utt was associated with ssS2 on the cell membrane. These observations indicate that the expressed S1 and S2 proteins associated in a similar manner to the authentic S1 and S2 proteins produced in DBT cells infected with cl-2. However, cell fusion was not observed in cells expressing either S1 or S2 nor in cells coexpressing both S1 and S2, although the whole S protein expressed by VV-TV did induce fusion.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">8245853</PMID><DateCompleted><Year>1993</Year><Month>12</Month><Day>27</Day></DateCompleted><DateRevised><Year>2020</Year><Month>04</Month><Day>15</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0022-1317</ISSN><JournalIssue CitedMedium="Print"><Volume>74 ( Pt 11)</Volume><PubDate><Year>1993</Year><Month>Nov</Month></PubDate></JournalIssue><Title>The Journal of general virology</Title><ISOAbbreviation>J. Gen. Virol.</ISOAbbreviation></Journal><ArticleTitle>Expression of the S1 and S2 subunits of murine coronavirus JHMV spike protein by a vaccinia virus transient expression system.</ArticleTitle><Pagination><MedlinePgn>2373-83</MedlinePgn></Pagination><Abstract><AbstractText>The spike (S) protein of murine coronavirus JHMV, variant cl-2, comprises two polypeptides, N-terminal S1 (with an N-terminal signal peptide) and C-terminal S2 (with a C-terminal transmembrane domain). In order to express these subunits, we constructed three different vaccinia virus transfer vectors (VV-TVs) containing cDNAs encoding the S1 protein without a transmembrane domain (pSFS1utt), the S1 protein with a C-terminal transmembrane domain derived from S2 (pSFS1tmd) or the S2 protein with an N-terminal signal peptide derived from S1 (pSFssS2). The S1 and S2 proteins were expressed in DBT cells by infection with vaccinia virus and transfection of these VV-TVs. In cells transfected with the pSFS1utt and pSFS1tmd, 96K and 106K proteins, respectively, were detected by Western blotting. The ssS2 protein expressed by pSFssS2 was 96K, which was slightly larger than the authentic S2 protein. The S1utt and S1tmd proteins were shown by binding studies using a panel of monoclonal antibodies to be antigenically indistinguishable from the authentic S1 protein. The S1tmd and ssS2 proteins were detected on the cell surface by immunofluorescence, whereas the S1utt protein was not. However, when the S1utt protein was expressed together with the ssS2 protein, the S1utt was detected on the cell membrane. This suggested that the S1utt was associated with ssS2 on the cell membrane. These observations indicate that the expressed S1 and S2 proteins associated in a similar manner to the authentic S1 and S2 proteins produced in DBT cells infected with cl-2. However, cell fusion was not observed in cells expressing either S1 or S2 nor in cells coexpressing both S1 and S2, although the whole S protein expressed by VV-TV did induce fusion.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Kubo</LastName><ForeName>H</ForeName><Initials>H</Initials><AffiliationInfo><Affiliation>National Institute of Neuroscience, NCNP, Tokyo, Japan.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Taguchi</LastName><ForeName>F</ForeName><Initials>F</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>England</Country><MedlineTA>J Gen Virol</MedlineTA><NlmUniqueID>0077340</NlmUniqueID><ISSNLinking>0022-1317</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D000956">Antigens, Viral</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="C578553">MHV surface projection glycoprotein</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D008562">Membrane Glycoproteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D011994">Recombinant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D064370">Spike Glycoprotein, Coronavirus</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D014759">Viral Envelope Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="C578557">spike glycoprotein, SARS-CoV</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D000595" MajorTopicYN="N">Amino Acid Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D000818" MajorTopicYN="N">Animals</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D000956" MajorTopicYN="N">Antigens, Viral</DescriptorName><QualifierName UI="Q000032" MajorTopicYN="N">analysis</QualifierName><QualifierName UI="Q000502" MajorTopicYN="N">physiology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D001483" MajorTopicYN="N">Base Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D015153" MajorTopicYN="N">Blotting, Western</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D002460" MajorTopicYN="N">Cell Line</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D005822" MajorTopicYN="N">Genetic Vectors</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008562" MajorTopicYN="Y">Membrane Glycoproteins</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D051379" MajorTopicYN="N">Mice</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008969" MajorTopicYN="N">Molecular Sequence Data</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D006517" MajorTopicYN="N">Murine hepatitis virus</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="Y">chemistry</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000502" MajorTopicYN="N">physiology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D011994" MajorTopicYN="N">Recombinant Proteins</DescriptorName><QualifierName UI="Q000096" MajorTopicYN="N">biosynthesis</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D064370" MajorTopicYN="N">Spike Glycoprotein, Coronavirus</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D014616" MajorTopicYN="N">Vaccinia virus</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D014759" MajorTopicYN="N">Viral Envelope Proteins</DescriptorName><QualifierName UI="Q000096" MajorTopicYN="N">biosynthesis</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000502" MajorTopicYN="Y">physiology</QualifierName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>1993</Year><Month>11</Month><Day>1</Day></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>1993</Year><Month>11</Month><Day>1</Day><Hour>0</Hour><Minute>1</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>1993</Year><Month>11</Month><Day>1</Day><Hour>0</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">8245853</ArticleId><ArticleId IdType="doi">10.1099/0022-1317-74-11-2373</ArticleId></ArticleIdList></PubmedData></pubmed></record>Pour manipuler ce document sous Unix (Dilib)
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